Microecology-turbidity toxin theory:correlation between helicobacter pylori infection and manifestation of tongue and gastroscopy

WANG Jin,WANG Huijie,XIAO Ying,GUO Jiaxuan,ZHAO Yubin

WANG Jin,Department of Integrated Chinese and Western Medicine,Hebei University of Chinese Medicine,Shijiazhuang 050000,China

WANG Huijie,the Traditional Chinese Medicine Hospital of Shijiazhuang Affiliated to Hebei University of Chinese Medicine,Shijiazhuang 050000,China

XIAO Ying,Department of Bioengineering,School of Chemical Engineering,Shijiazhuang University,Shijiazhuang 050000,China

GUO Jiaxuan,The Traditional Chinese Medicine Hospital of Shijiazhuang Affiliated to Hebei University of Chinese Medicine,Shijiazhuang 050000,Hebei,China

ZHAO Yubin,Department of Integrated Chinese and Western Medicine,Hebei University of Chinese Medicine,Shijiazhuang,China;Shijiazhuang People's Hospital,Shijiazhuang 050000,China;the Traditional Chinese Medicine Hospital of Shijiazhuang Affiliated to Hebei University of Chinese Medicine,Shijiazhuang 050000,China

Abstract OBJECTIVE:To clarify the relationship between helicobacter pylori (Hp) infection,tongue manifestation and gastroscopic manifestation,and explore the mechanism of action of“oral-gastric microecology”in the occurrence and development of stomach-related diseases.MEDHODS:A total of 1100 patients were divided into Hp positive group and Hp negative group according to 13C breath test results.The tongue manifestation materials from patients were collected to make tongue printing slides for morphological observation of exfoliative cells from tongue coating,and electronic gastroscopy was performed on the patients and the data was analyzed.RESULTS:Hp infection rate was about 45.95%;and the greasy coating was mostly observed in Hp positive group(63.14%,P ＜ 0.05).The total tongue coating distribution was as follows:yellow greasy coating (56.66%, P ＜0.05) ＞ yellow coating ＞ white coating ＞ white greasy coating.The gastric mucosal roughness and mucosal hematoplastic plaques in the two groups were more common in the Hp positive group (P ＜ 0.05).There was a significant difference in the granular subgroup and in the rough mucosal group (P ＜ 0.05).CONCLUSIONS:Hp infection is more commonly observed in yellow greasy coating,and this can be used as a reference for Traditional Chinese Medicine in determining whether it is Hp infection or not.Mucosal granule roughness,hyperemia,and oral Hp showed association with Hp infection in the stomach and interacted with micro-ecological environment that is composed of the oral cavity.

Keywords:Helicobacter pylori;abnormal tongue presentations;gastroscopy;breath tests;micro-ecology;turbidity toxin theory

1.INTRODUCTION

Helicobacter pylori (Hp) is recognized as a pathogenic factor for stomach-related diseases.Hp is closely related to gastrointestinal diseases such as chronic gastritis,peptic ulcer,gastric mucosa-associated lymphoid tissue lymphoma,and gastric cancer,and therefore,it is included in class I carcinogens.1In recent years,increased attention has been paid to Hp infections and their corresponding therapeutic drugs.This is because the Hp infection rate in most of the countries shows a downward trend,but still remains to be high.2,3

Tongue manifestation can objectively reflect the depth of the disease location,nature of the pathogenic factors,and the rise and the fall of vitalQiand pathogen and disease tendency,which forms an important basis for clinical treatment based on syndrome differentiation.4

Tongue coating can be used for diagnosing conditions of 5 internal organs.The turbidity toxin theory of traditional Chinese medical science is an integral part of the academic theoretical system of Traditional Chinese Medicine (TCM).The theory of“microecology-turbidity toxin”is a further elaboration of the pathogenic mechanism of turbidity toxin and the changes of pathogenesis when the human body is in a state of turbidity toxin.5Hp,as a pathogen that invades the human body,belongs to the category of“toxic pathogens”in turbidity toxin theory of TCM,and is also an important factor that initiates the occurrence of gastric-related diseases.Therefore,the pathological state caused by Hp can reflect in the tongue manifestation.

With the development of medical examination methods,TCM has been moved from macroscopic syndrome differentiation to microscopic syndrome differentiation.Objectification and standardization are the important directions for the development of TCM.This study with the guidance of“microecology-turbidity toxin”theory found the relationship between Hp infection and tongue manifestation,the correlation between Hp infection and tongue manifestation and gastroscope manifestation was clarified with gastroscope manifestation as the entry point,and the mechanism of“Oral-gastric microecology”in the occurrence and development of gastric diseases was explored to provide data for clinical diagnosis and treatment.

2.MATERIALS AND METHODS

2.1.Clinical data

From August 2015 to October 2017,patients who underwent gastroscopy in the endoscopy center of our hospital met the following inclusion criteria:

Patients (a) with age range from 18 to 75 years;(b) with symptoms of upper digestive system;(c) without Hp radical treatment within four weeks;(d) who provided signed informed consent and was willing to cooperate with the study.

The exclusion criteria were as follows:(a) patients with severe cardiovascular and cerebrovascular diseases,liver and kidney diseases,etc.;(b) patients with diseases that could change the tongue manifestation significantly,such as acute upper respiratory tract infection;(c)patients who had taken proton pump inhibitors or bismuth and other drugs in the previous 4 weeks;(d)pregnant and lactating women;and (e) patients with a history of nervous system and mental diseases.

2.2.Research method

Research contents and observation index:(a) the general information of patients;(b) the main symptoms of upper gastrointestinal diseases were collected;(c) the information of whether the patient had taken relevant TCM within half a year was analyzed;(d) tongue manifestation pictures of the patient and the related indicators of tongue manifestation were collected;(e)morphological observation indexes of tongue coating exfoliated cells such as maturity index (MI) and maturity value (MV);and (f) gastroscopic appearance.The patients who met the inclusion criteria should fill in the Patient Information Observation Table.Tongue manifestation pictures and exfoliated cells of tongue coating were taken,gastroscopy was performed under endoscopy to observe and detect whether they were infected with Hp,and the observed data of the study subjects were sorted out and then recorded.

This study was approved by the Ethics Committee of the Hospital,and all patients signed the informed consent form.

2.3.Collection method

Patients were fasted in the morning without gargling.The tongue was naturally extended under the same light intensity before undergoing gastroscopy,and the image of the tongue was taken by the same digital camera.

2.4.Diagnosis standard for tongue manifestation

The tongue manifestation analysis software (Intelligent Auxiliary Diagnosis System for Tongue Manifestation V1.0) was used to analyze the collected tongue manifestation pictures.The diagnosis was made by 2 senior traditional Chinese physicians by referring to theDiagnostics of Chinese Medicine.6Their diagnoses were compared with the results of the analysis software.With the diagnostic results,tongue manifestation diagnosis of patients could be concluded.

Instrument:(a) Tongue manifestation acquisition rack(Shijiazhuang,China),patent number:2017201 294764;(b) Canon EOS 550D SLR digital camera (zoom lens EFS 18-55mm f/3.5-5.6 IS) (Beijing,China).

2.5.Morphological observation on exfoliative cells of tongue coating-tongue printing slide

Experiment materials:(a) reagent:Papanicolaou staining solution,purchased from Zhuhai Baso Biotechnology Co.,Ltd.(Zhuhai,China),production batch number:716091.

(b) Instrument:optical microscope:OLYMPUS BX43F(Beijing,China).Combination dyeing cylinder,cover glass slide,glass slide,etc.

The methods of preparation,fixation,staining and observation of tongue print were judged according to relevant literature.7

Morphological test criteria for exfoliated cells of tongue coating were judged according to relevant literatures.8

2.6.Collection and observation indexes of gastroscopy manifestation

FUJINON XL4450 electronic gastroscope-600WR was used for observation and indexes were referred to relevant literatures and guidelines.10

2.7.Hp examination

Experimental materials:the reagent of13C breath test pill box (Beijing Younite Pharmaceutical Co.,Ltd.(Beijing,China),production batch number:21161101) was used.The instrument of WLD600C13C breath analyzer(Beijing Wanliandaxinke Instruments Co.,Ltd.,Beijing,China) was used.The urea breath test was used and the test method done according to the manufacturer’s instructions.

According to the DOB value results,Hp infection was judged to be positive if the DOB value was greater than 4.0,and negative if the DOB value was less than 4.0.

2.8.Data processing and statistical methods

Excel software was used to build the database,and statistical software STATA 14.2 (TX,USA) was used for statistical processing.χ2test of R × C table was used for patient observation index data,and Fisher’s exact probability method was used for statistical analysis of some indicators.Two independent samples were analyzed by approximatet-test and subjected to multiple logic regression.Statistical significance was set atP＜0.05.

3.RESULTS

A total of 1117 patient information observation tables were distributed and recovered.Seventeen observation tables were not included in the research due to inadequate data filling,and so 1100 patients were included in the research.

3.1.Basic conditions of patients

Among the 1100 patients,659 (59.91%) were females and 441 (40.09%) were males.The average age of the patients was (49 ± 13) years,and there were 421 young people (＜ 45 years,38.27%),410 middle-aged people(45-59 years,37.27%),and 269 old people (more than 60 years,24.46%).There were 510 cases (45.95%) in the Hp positive group.The negative Hp infection group included 590 patients (53.15%).

3.2.General information of patients and factors related to Hp infection

Hp infection was defined as a dependent variable,gender,age,medication status,smoking and drinking history in the recent period were included as independent variables.The results showed no significant differences in the number of cases (P＞ 0.05).

Hp infection was defined as an independent variable,and the correlation between Hp infection and tongue manifestation changes was judged by analyzing tongue manifestation of the patients.

3.3.Relationship with tongue body

The distribution of tongue color in Hp-positive cases was mostly light red,followed by crimson,red,and finally blue and purple tongue.There were a total of 301(59.01%) patients with light red tongue,70 (13.73%)patients with red tongue,91 (17.84%) patients with crimson tongue and 48 (9.41%) patients with blue and purple tongue.After χ2analysis,the results showed statistically significant differences were observed (P=0.008).

The results showed that the tongue shape (tough tongue,tender tongue,spotted tongue,teeth-marked tongue and fissured tongue) had no clear relationship with Hp infection.The color coat of Hp positive cases was more yellow than white.Among them,there were 419 (82.16%)patients with yellow coating and 91 (17.84%) patients with white coating,which indicated that the number of yellow coating cases was obviously higher than the number of white coating cases.After χ2test,the results showed statistically significant differences (P=0.000).The results showed no clear relationship between the thickness and stripping of coating texture and Hp infection.Greasy coating in Hp positive cases was more than those with greasy-free coating,and there were 322(63.14%) patients with greasy coating and 188 (36.86%)patients with greasy-free coating.This indicated that the number of greasy coating cases was higher than that of greasy-free coating cases.After χ2test,the results showed statistically significant differences (P=0.015).After integration analysis of tongue color and greasy tongue,the distribution of coating in Hp-positive cases was mostly yellow,followed by yellow,white,and finally white greasy coating.Among them,289 cases of yellow greasy coating accounted for 56.66%;41 cases of white greasy coating accounted for 8.04%;130 cases of yellow coating accounted for 26.53%;and 50 cases of white coating,accounting for 10.20%.The number of cases of yellow greasy coating was more than that of other tongue coating cases.After Fisher's exact probability analysis,the results showed statistically significant differences (P=0.000).

3.4.Correlation between Hp infection and morphology of exfoliated cells of tongue coating

MI of basal layer cells:both the Hp positive group and the Hp negative group were 0.MI of intermediate cells:the positive group of Hp infection was 24 ± 9,and the negative group of Hp infection was 19 ± 10.MI of surface cells:the positive group with Hp infection was 76 ± 9,and the negative group with Hp infection was 81± 10.MV of exfoliated cells of tongue coating:the positive group of Hp infection was 88 ± 4,and the negative group of Hp infection was 90 ± 5.After approximatet-test on two independent samples,the results showed statistically significant differences (P=0.0181).

3.5.Relationship between Hp infection and gastroscopy changes

Hp infection was taken as independent variable and gastroscopy manifestation of patients was taken as dependent variable,and the correlation between the two was observed.

The results showed that gastric mucosa color,mucous erosion,mucous bleeding spots,mucous hyperemia spot,polyps,yellow pigment tumor,and appearance of mucosal vascular transmission demonstrated no definite relationship with Hp infection.

The distribution of rough gastric mucosa in Hp positive infected cases was mostly granular,followed by roughfree,and finally nodular,in which there were 189(37.05%) rough-free cases,71 (13.92%) nodular cases,and 252 (49.41%) granular cases (P=0.000).After the disordered multi-classification logistic regression analysis (Table 6),the gastric mucosa of the granular and nodular rough groups was comparable with that of rough-free mucous group,and the results showed no statistical significance in the nodular group (P=0.069),while statistically significant in the granular group (P=0.000).

4.DISCUSSION

4.1.Hp infection

China has a high Hp infection rate in the world.10The Hp Research Cooperation Group of China11investigated the Hp infection rate in the general population of 19 provinces,cities and autonomous regions,which showed a total infection rate of 56.22%,and the infection rate of Hebei Province is 45.39%,which is basically the same as the results put forwarded in our study (45.95%).

4.2.Correlation between Hp infection and tongue manifestation

Tongue manifestation includes tongue body and tongue coating.Tongue body indicates the prosperity ofQiand the blood of internal organs,for which it can be used for diagnosing as weak and strong points of the internal organs,prosperity and declination ofQiand blood.Another study of ours identified the influences ofHp infection on gastric and tongue coating microbial populations.Infection with CagA+Hp strains reduced the diversity of gastric microbiota and altered its composition and functions.6However,as an external pathogen invading the body,Hp is more closely related to tongue coating,and the changes in tongue body quality reflects the damage caused by Hp to the body.

4.3.Tongue color

Tongue color indicates weak or strong points of internal organs and prosperity ofQiand blood.6With regard to the relationship between Hp and tongue color,the results of our study showed that the majority of Hp infected patients had light red tongue,which is the same as the results of other studies.12This suggests that the tongue color of patients with Hp infection is the same as that of normal people.This is due to the fact that short time from the invasion of external pathogenic factor into the body or weak pathogenicity (such as II type low toxin strain infection) or no damage to healthyQiare not enough for causing the losses ofQi-blood and body fluids,further causing the change of tongue color.In this research,most of the gastroscopy images of the patients are light in color under endoscopy,showing that Hp’s invasion of the body has not caused a large change in the body,and so the tongue color still remains to be normal.This was consistent with the saying by traditional Chinese medical science that a light red tongue can be seen in healthy people,external contraction and exterior pattern,internal injuries and mild disease.6Therefore,it is not appropriate to use light red tongue as a key screening index for Hp infection.This research showed no significant difference between Hp and tongue shape (P＞ 0.05),which is due to the fact that under the influence of various factors,short time since the invasion of external pathogenic factor in the body or weak pathogenicity or no damage to healthyQiare considered not enough to cause the loss ofQiblood and body fluids.This was consistent with the conclusion that light red tongue was commonly observed during Hp infection.

4.4.Tongue coating

The tongue coating consists of two parts,coating body and coating color.This study showed that 698 Hp infected patients had yellow coating,accounting for 63.64% (P ＜0.05).As Hp infection spreads to the gastric mucosa through the digestive tract,it causes certain damage to the gastric mucosa.The damage should involve the evidence of internal injuries,and so the Hppositive patients mostly have yellow coating.Research on the correlation between Hp and coating body showed:(a) no statistically significant difference between Hp and the thickness of coating and peeling coating,and this might be related to the influence of different courses of disease or pathogenicity of Hp with regard to the formation of thick or thin coating.As the peeled coating is rarely seen in general population,it had a certain effect on the results.(b) A statistically significant difference between Hp and greasy coating was observed (P ＜0.05),which was similar to that of temperature results.13The statistically significant coating color and greasy coating are integrated and analyzed,and most of the cases had yellow greasy coating,followed by yellow coating,white coating,and finally the white greasy coating (P ＜0.05),in which 289 cases had yellow greasy coating,which accounted for 56.66%,and this was similar to the results of other studies.14Hp infection is more common in the greasy coating and yellow greasy cases,which might be due to Hp invasion into the human body,and implanting into the gastric mucosa,which causes damage to the physiological function of the internal organs and abnormal transport of spleen and stomach,interior accumulation of damp-turbidity and phlegm-rheum overflowing and staying on the tongue surfaces,or persistent damp-turbidity transforming into heat.

4.5.Exfoliated cells of tongue coating

The main basis for the formation of tongue coating involves normal proliferation,migration and keratinization of epithelial cells of the tongue.The components mainly include keratinized trees with filamentous papilla differentiation and exfoliated cells,bacteria,etc.15Tongue printing method is used to observe the composition of exfoliated cells of the tongue,and the observation indexes included MI and MV.The effect of Hp infection on tongue coating involves studying from the microscopic level,which supplement the deficiency of naked eye observation of tongue coating.This research indicates that the maturity of exfoliated cells on tongue coating in Hp infected patients was lower than those in Hp infection-free patients,and Hp infection affects the maturation of exfoliated cells of tongue coating.

4.6.Proposal of“oval cavity-stomach ecology symbiosis chain”based on“micro-ecology-turbidity toxin”theory

According to the“micro-ecology-turbidity toxin”theory,5the microbial communities of oral cavity and gastrointestinal tract are interrelated and interact with each other,and the oral cavity and stomach together constitute the ecological microenvironment of microbial symbiosis in the human body.When the human body that acts as a host is healthy,then the normal flora of the oral cavity and the stomach together resists the colonization of external bacteria and stimulates the host’s immune system,which has a positive effect on the host’s immune regulation.The human body provides living environment for microorganisms and the nutrients needed for growth and proliferation of microorganisms.The normal microorganisms in the oral and gastric mucosa are present in dynamic balance.If the body is in a pathological condition,then an abnormal growth of microorganisms occurs in the oral cavity and stomach,leading to invasion of foreign microbes and ultimately destroying the balance of the micro-ecological environment.

In the oral cavity-stomach micro-ecological system,tongue coating acts as a sensitive indicator of the overall micro-ecological balance.From the point of view of TCM,tongue is a sign that reflects the spleen transport function,and tongue coating is the main reflection of tongue surface after the stomachQidigesting water and cereal.When the human body is in a healthy state,then the tongue appears as a shallow white tongue coating,and the micro-ecology of the tongue coating is in a state of dynamic equilibrium.In this micro ecosystem,the number of microorganisms remains to be constant,and this is referred to as the state of“Yin-Yangbalance”.If the body is affected by the pathological factors,then the functions of internal organs are changed accordingly and the micro-ecological balance of the tongue coating is broken,causing abnormal microbe species and quantity of the tongue coating,further leading to pathological coatings.Based on this,pathological change of human tongue coating occurs due to the change of oral microecological environment,which is highly consistent with the theory of TCM.

In the future,methods such as magnifying endoscopy and staining should be used to determine gastroscopy manifestations in order to make the results more accurate.The microbiology and gene level analysis on the tongue coating should be carried out to further reveal the essence of the changes in tongue manifestation and gastroscopy manifestation caused by Hp infection,and to establish objective diagnosis standard for traditional Chinese medical science in determining Hp infection during inspection diagnosis on tongue manifestation and gastroscopy manifestation.

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